An improved understanding of cancer immunology has allowed significant advances in clinical translation. Dramatic clinical responses have been observed by other groups and ours with adoptive cell transfer therapy using T cell receptor (TCR)-engineered T lymphocytes in patients with metastatic melanoma. These response rates exceed 50%, however, in our experience, patients eventually relapse. We hypothesize that these highly activated and cytotoxic T lymphocytes, when entering an immunosuppressive tumor microenvironment, lose their effector function over time. Tumor secreted TGF is a major contributor to the immunosuppressive properties of the tumor microenvironment. We hypothesize that incorporating a dominant-negative TGF receptor II (dnTGFRII) into the MART-1 TCR retroviral vector used to transduce patient T lymphocytes will render these cells insensitive to TGF, resulting in an enhancement of their effector function in the melanoma tumor microenvironment. We propose to test this hypothesis in a clinical investigation of patients with metastatic melanoma in which two populations of MART-1 TCR transgenic T cells are co-infused: one population retrovirally transduced with MART-1 TCR and the other population transduced with the same TCR and the dnTGFRII. The first vector (GMP-grade) is in hand, and the second vector, already pre-clinically tested, will be manufactured at the Indiana University National Viral Production Facility (IU VPF). These two populations of engineered T cells will be reintroduced into the patient after a lymphodepleting conditioning regimen. The adoptively transferred cells will be supported in vivo by systemic administration of interleukin-2 and MART-126-35 peptide-pulsed dendritic cell vaccines. In this phase I clinical trial, the primary end-points are safety and feasibility; the secondary end-points are the study of differential persistence of the two transgenic cell populations, and the analysis of their key immunological signatures; and the tertiary end point will be clinical response. Our specific aims are: 1) To evaluate, in a phase I trial, the safety, persistence and clinical response of adoptive transfer of MART-1 TCR/dnTGFRII engineered T cells in metastatic melanoma patients. 2) To evaluate key immunological end-points that correlate with the degree of clinical responses. We will collect, at defined time points, and analyze peripheral blood and tumor infiltrating lymphocytes from biopsies for number, immunophenotyping, and functional assays. Our group at UCLA, with a MART-1 TCR clinical trial experience, has learned that a critical issue is the lack of long-term specific T cell persistence and function. This present proposal will directly address this problem with the goal of achieving sustained clinical responses.